The unifying hypothesis of this proposal is that macromolecular complex of type 2 lysophosphatidic acid receptor (LPA2 receptor) and Na+/H+ exchange regulatory factor-2 (NHERF2) plays important roles in the pathogenic process of secretory diarrhea. We aim to study the formation and regulation of the macromolecular complex at the molecular level under physiological and pathophysiological conditions that play a critical role in cholera induced diarrhea. Moreover, the research will study the mechanism through which LPA2 receptor- mediated signaling events regulate CFTR-dependent electrolyte secretion in cells and fluid secretion in an animal model of diarrhea. Three aims will be studied: Aim 1: To test the hypothesis that LPA2 receptor is primarily expressed on the luminal surface of the gut and is down regulated in certain forms of diarrheal diseases, and to test the hypothesis that activation of LPA2 receptor inhibits compartmentalized cAMP generation at the plasma membrane. Aim 2: To test the hypothesis that LPA can regulate the formation of the LPA2-containing macromolecular complex by further clustering it to microdomain on the plasma membrane and that the process is mediated by NHERF2-dependent protein-protein interactions Aim 3: To test the hypothesis that disruption of the macromolecular complex by silencing NHERF2 alters compartmentalized cAMP levels at the plasma membrane and that in vivo NHERF2 knockout mice generate altered cAMP in response to CTX. The proposal is of great importance and significance for understanding the pathogenic process of the deadly secretory diarrheal diseases at the molecular level, and for possible therapeutic intervention of the disease. The proposed study is highly innovative in both conceptual advance and in technology/technique development. PUBLIC HEALTH RELEVANCE: In this study we propose that the macromolecular complex of LPA2 and NHERF2 is regulated by LPA signaling, which further clusters the macromolecular complex to subdomain on the plasma membrane, and that the macromolecular complex plays important role in regulating compartmentalized cAMP level in proximity to CFTR channel, thus regulating CFTR-dependent electrolyte and fluid secretion in gut epithelial cells and in the mouse model. Therefore, the macromolecular complex plays an important role in pathogenic process of secretory diarrhea. By targeting the macromolecular complex, we may find ways to control fluid secretion and thereby control or cure the diarrheal disease.